1. Field of the Invention
The present invention relates to a process for recovering protease. More particularly, it relates to a process for recovering protease by isolating the protease from an addition compound after a peptide synthesis of peptide-bonding amino acid derivatives in the presence of the protease.
2. Description of Prior Arts
It has been known that proteases such as papain and chymotrypsin are used for forming peptide bonds as the reverse reaction of protein decomposition. For example, anilides have been produced by using papain by Bergman and the peptide syntheses using monoamino-carboxylic acids such as leucine having N-terminal protective benzoyl group and leucine and glycine having C-terminal protective amide or anilide group have been attained with papain and chymotrypsin by Fruton. (Advances in Protein Chemistry Vol. 5, page 33 (1949). Academic Press Inc. New York, N.Y.).
Recently, some of the inventors Isowa et al. reported peptide syntheses using amino acids having an N-terminal protective benzyloxycarbonyl group and amino acids having a C-terminal ester group with enzymes such as papain, Prolisin, Subtilisin BPN' etc. (Nippon Kagakukai 35th Autumn Meeting Brief Report Pages 482 and 486 (1976) Nippon Kagakukai).
Some of the inventors and Isowa et al. proposed the reaction as asparatic acid or glutamic acid having N-terminal protective group with a monoaminomonocarboxylic acid having C-terminal protective group which has no other functional group in the presence of protease whereby an addition compound of the monoaminomonocarboxylic ester having no other functional group and the reaction product was formed, and the isolation of the addition product was carried out. (Japanese Patent Application No. 7279/1977; U.S. Pat. application No. 870,108; Canadian Pat. application No. 295,711 and West German Patent Application P 2801238.6).
The protease used in the process is considered to preserve the protease activity after the reaction. However, the filtrate of the reaction mixture from which the reaction product is separated (The reaction product is deposited to be insoluble in the medium for the reaction.) has only weak protease activity.
According to the studies of the inventors, it has been found that most of the protease used in the reaction is separated together with the reaction product from the filtrate and preserves the protease activity and can be reused for the peptide synthesis etc. after isolating the protease from the reaction mixture.